Abstract
The colonic epithelium plays a crucial role in gastrointestinal homeostasis, and colon organoids enable investigation into the molecular mechanisms underlying colonic physiology. However, the method for differentiating induced pluripotent stem cells (iPSCs) into human colon organoids (HCOs) is not necessarily standardized, and studies using HCOs are limited. This study refines the differentiation of HCOs by comparing two protocols reported in Cell Stem Cell and Nature Medicine journals. The former protocol, which uses transient bone morphogenetic protein 2 (BMP2) signaling activation, demonstrated superior efficacy in upregulating colon-specific markers. Additionally, adenovirus-mediated transduction of the transcription factors HOXD13 or SATB2 during hindgut endoderm development, together with BMP2 treatment, enhanced colonic identity, suggesting improved colonic maturation. This optimized protocol advances the generation of mature HCOs, offering a better model for investigating colonic epithelial biology and pathology.